Highly efficient production of an influenza H9N2 vaccine using MDCK suspension cells

Yixiao Wu,Wen-Song Tan,Hanjing Jia,Hanzhang Lai,Xuping Liu

doi:10.1186/s40643-020-00352-4

Abstract

The use of H9N2 subtype avian influenza vaccines is an effective approach for the control of the virus spread among the poultry, and for the upgrading of vaccine manufacturing, cell culture-based production platform could overcome the limitations of conventional egg-based platform and alternate it. The development of serum-free suspension cell culture could allow even higher virus productivity, where a suspension cell line with good performance and proper culture strategies are required. In this work, an adherent Mardin–Darby canine kidney (MDCK) cell line was adapted to suspension growth to cell concentration up to 12 × 106 cells/mL in a serum-free medium in batch cultures. Subsequently, the H9N2 influenza virus propagation in this MDCK cell line was evaluated with the optimization of infection conditions in terms of MOI and cell concentration for infection. Furthermore, various feed strategies were tested in the infection phase for improved virus titer and a maximum hemagglutinin titer of 13 log2 (HAU/50 μL) was obtained using the 1:2 medium dilution strategy. The evaluation of MDCK cell growth and H9N2 virus production in bioreactors with optimized operating conditions showed comparable cell performance and virus yield compared to shake flasks, with a high cell-specific virus yield above 13,000 virions/cell. With the purified H9N2 virus harvested from the bioreactors, the MDCK cell-derived vaccine was able to induce high titers of neutralizing antibodies in chickens. Overall, the results demonstrate the promising application of the highly efficient MDCK cell-based production platform for the avian influenza vaccine manufacturing.

Highlights

IntroductionThe H9N2 viruses have been isolated globally in the past few decades and are persistently circulating in several countries in Asia, the Middle East, and North Africa (Gu et al 2017)
Avian influenza virus (AIV) can cause infections both in animals and humans
Advances in the medium development facilitate an easier and efficient cell adaptation to growth in suspension, and for the first time, a fast adaptation of an adherent Mardin–Darby canine kidney (MDCK) cell line to grow in single suspension in a serumfree medium after only 19 days was demonstrated

Summary

Introduction

The H9N2 viruses have been isolated globally in the past few decades and are persistently circulating in several countries in Asia, the Middle East, and North Africa (Gu et al 2017) This has resulted in severe economic burdens to the poultry industry by the decrease in egg production and the moderate-to-high mortality of poultry (Lamb and Takeda 2001; Lee and Song 2013; Pu et al 2015). The conventional embryonated egg production platform has been intensively applied for the manufacturing of influenza vaccines for more than 70 years. This platform is highly dependent on the supplies of eggs, which can be limited in the event of an influenza pandemic (Hegde 2015; Ulmer et al 2006). By using the MDCK cells, influenza vaccines, such as F lumist® (MedImmune), Flucelvax®/Optaflu® (Seqirus/Novartis), and S KYCellflu® (SK chemicals), have been developed and certified (Genzel and Reichl 2009; Sun et al 2011)

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Conclusion